PREFACE
Dr. Sheldon Wolf and his colleagues in the USA proposed the use of the term "radioadaptive response" about 1980. A priming irradiation with a low dose or delivered at a low dose rate was observed to depress the biological effects induced by a subsequent high dose irradiation. This phenomenon had already been observed after exposures to UV light and to heat. UV-reactivation had been described, and this meant that UV-irradiated phage had higher survival rates in Escherichia coli host cells which had been irradiated with a low dose of UV light prior to infection. Lower survival rates were seen for phage in host cells which had not been exposed to UV prior to infection. In the case of heat, cells exposed to a mild heat treatment acquired thermotolerance against subsequent severe heat exposures. The development of this thermotolerance was shown to depend on the induction of heat shock proteins. Therefore, when the first reports of the radioadaptive response were made, scientists were already aware of such a phenomenon. In addition, this adaptive behavior can be considered quite similar to the immune response which is initiated by a weak exposure to an antigen.
Organisms on the earth exist and adapt to environmental changes through the aid of many types of biological functions. There is currently no answer to the question of whether many different organisms can adapt and survive by using the same adaptive mechanisms. However, this is a very interesting area for science to focus on. In this special issue, recent papers are presented discussing the radioadaptive response. These papers were presented in a workshop at the annual meeting of the Japanese Radiation Research Society (Dr. T. Norimura, president) in Kitakyushu City in November, 2008. I hope these manuscripts will provide useful and helpful information for those interested in current and future progress in the study of the radioadaptive response.

Translated and modified from Radiat. Biol. Res. Comm. Vol.43(4); 408, (2008, in Japanese).

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Viable organisms recognize and respond to environmental changes or stresses. When these environmental changes and their responses by organisms are extreme, they can limit viability. However, organisms can adapt to these different stresses by utilizing different possible responses via signal transduction pathways when the stress is not lethal. In particular, prior mild stresses can provide some aid to prepare organisms for subsequent more severe stresses. These adjustments or adaptations for future stresses have been called adaptive responses. These responses are present in bacteria, plants and animals. The following review covers recent research which can help describe or postulate possible mechanisms which may be active in producing adaptive responses to radiation, ultraviolet light, and heat.

Translated and modified from Radiat. Biol. Res. Comm. Vol.43(4); 409-423, (2008, in Japanese).

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We studied cellular responses in normal human fibroblasts induced with low-dose (rate) or low-fluence irradiations of different radiation types, such as gamma rays, neutrons and high linear energy transfer (LET) heavy ions. The cells were pretreated with low-dose (rate) or low-fluence irradiations (~1 mGy/7-8 h) of ¹³⁷Cs gamma rays, ²⁴¹Am-Be neutrons, helium, carbon and iron ions before irradiations with an X-ray challenging dose (1.5 Gy). Helium (LET = 2.3 keV/μm), carbon (LET = 13.3 keV/μm) and iron (LET = 200 keV/μm) ions were produced by the Heavy Ion Medical Accelerator in Chiba (HIMAC), Japan. No difference in cell-killing effect, measured by a colony forming assay, was observed among the pretreatment with different radiation types. In mutation induction, which was detected in the hypoxanthine-guanine phosphoribosyltransferase (hprt) locus to measure 6-thioguanine resistant clones, there was no difference in mutation frequency induced by the X-ray challenging dose between unpretreated and gamma-ray pretreated cells. In the case of the pretreatment of heavy ions, X-ray-induced mutation was around 1.8 times higher in helium-ion pretreated and 4.0 times higher in carbon-ion pretreated cells than in unpretreated cells (X-ray challenging dose alone). However, the mutation frequency in cells pretreated with iron ions was the same level as either unpretreated or gamma-ray pretreated cells. In contrast, it was reduced at 0.15 times in cells pretreated with neutrons when compared to unpretreated cells. The results show that cellular responses caused by the influence of hprt mutation induced in cells pretreated with low-dose-rate or low-fluence irradiations of different radiation types were radiation-quality dependent manner.

Translated and modified from Radiat. Biol. Res. Comm. Vol.43(4); 424-435, (2008, in Japanese).

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The biological effects of low-dose radiation are not only of social concern but also of scientific interest. The radioadaptive response, which is defined as an increased radioresistance by prior exposure to low-dose radiation, has been extensively studied both in vitro and in vivo. Here we briefly review the radioadaptive response with respect to mutagenesis, survival rate, and carcinogenesis in vivo, and introduce our recent findings of cross adaptation in mouse thymic cells, that is, the suppressive effect of repeated low-dose radiation on mutation induction by the alkylating agent N-ethyl-N-nitrosourea.

Translated and modified from Radiat. Biol. Res. Comm. Vol.43(4); 436-442, (2008, in Japanese).

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In this review, we would like to introduce a unique approach for the estimation of radioadaptation. Recently, we proposed a new methodology for evaluating the repair efficiency of DNA double-strand breaks (DSB) using a model system. The model system can trace the fate of a single DSB, which is introduced within intron 4 of the TK gene on chromosome 17 in human lymphoblastoid TK6 cells by the expression of restriction enzyme I-SceI. This methodology was first applied to examine whether repair of the DSB (at the I-SceI site) can be influenced by low-dose, low-dose rate gamma-ray irradiation. We found that such low-dose IR exposure could enhance the activity of DSB repair through homologous recombination (HR). HR activity was also enhanced due to the pre-IR irradiation under the established conditions for radioadaptation (50 mGy X-ray-6 h-I-SceI treatment). Therefore, radioadaptation might account for the reduced frequency of homozygous loss of heterozygosity (LOH) events observed in our previous experiment (50 mGy X-ray-6 h-2 Gy X-ray). We suggest that the present evaluation of DSB repair using this I-SceI system, may contribute to our overall understanding of radioadaptation.

Translated and modified from Radiat. Biol. Res. Comm. Vol.43(4); 443-453, (2008, in Japanese).

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Exposure to low doses of radiation has been recently proven to be much more mutagenic and carcinogenic than previously thought. Since radiation sensitivity varies with different phases of cell cycle, we have investigated the activation of protein kinase C (PKC) after low doses (0.10-1 Gy) of γ-irradiation on proliferating (log) and non-proliferating (confluent/plateau) human normal lung fibroblast (MRC-5) cells. PKC isoforms have been shown to play key roles in the regulation of proliferation, differentiation, migration and survival. In this study, we have examined the activation of phosphorylated forms of PKC isoforms (PKC-βII, PKC-α/β, PKC-θ) and non-phosphorylated PKC-α in an attempt to understand its kinases in total and subcellular (cytosolic and nuclear) fractions. Cytosolic fraction of the log phase cells showed an increase in activity of PKC-βII, PKC-α/β and PKC-θ with the radiation dose. However, in the nuclear fraction, PKC-βII and PKC-θ showed higher activity than the PKC-α/β. In the plateau phase cells of the cytosolic fraction, PKC-βII showed higher activity than the PKC-α/β and PKC-θ isoforms. Furthermore, in the nuclear fraction PKC-βII and PKC-α/β isoforms showed higher activity than the PKC-θ. In total cellular protein of the log phase cells, a dose dependent increase in PKC-βII activity followed by PKC-α/ β was observed and in the plateau phase of cells, PKC-βII showed higher activity than the PKC-α/ β. The specific activation of PKC isoforms in the plateau phase cells, as demonstrated for the first time, may help us to understand the radiation induced initiation of cellular transformation like hyper-proliferative phenotype, if any.

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Two extracts E1and E2 were prepared from the dried root of the plant Caesalpinia digyna by extracting with solvents of different polarity. The extracts were standardized with respect to a polyphenol, bergenin, by LC- MS analysis and they were subjected to free radical scavenging activity and in vitro radioprotection studies. Free radical reactions were carried out with superoxide, hydroxyl, and peroxyl radicals and DPPH. In vitro radioprotecting activity was studied by following their effect on γ-radiation induced lipid peroxidation, protein carbonylation and DNA damage. The results indicated that E1 with higher free radical scavenging ability is also a more potent inhibitor of radiation induced damage to proteins, DNA and liposomes than E2. Comparing the results with those for bergenin indicated that bergenin alone is not responsible for the free radical scavenging ability and in vitro radioprotection. The studies also confirmed that the extracts enriched with bergenin are more effective than the isolated polyphenol, bergenin.

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Proton magnetic resonance spectroscopy (¹H MRS) is a potential method to detect and quantify a boron neutron capture therapy ¹⁰B-carrier compound, L-p-boronophenylalanine (BPA), in the brain. However, optimal positioning of MRS voxel to capture tissue with maximal BPA concentration can be challenging. Three dimensional proton magnetic resonance spectroscopic imaging (3D ¹H MRSI) provides spectral data covering a large spatial volume, which is a major advantage in detecting and quantifying BPA. BPA detection limit in phantom conditions was determined at 1.5 T using a 3D ¹H MRSI protocol with clinically acceptable nominal spatial resolution and duration. Quantification tests for aqueous phantom were performed using both single voxel MRS and 3D MRSI. In 3D MRSI, BPA detection limit was ~1.0 mM and BPA quantification accuracy was better than ±5%. The results suggest that MRSI would be a feasible method for in vivo BPA evaluation in clinical conditions.

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To study mechanisms which could be involved in the reverse dose rate effect observed during mutation induction after exposure to high LET radiation, synchronized mouse L5178Y cells were exposed to carbon 290 MeV/n beams with different LET values at the G2/M, G1, G1/S or S phases in the cell cycle. The frequency of Hprt-deficient (6-thioguanine-resistant) mutant induction was subsequently determined. The results showed that after exposure to high LET value radiation (50.8 and 76.5 keV/μm), maximum mutation frequencies were seen at the G2/M phase, but after exposure to lower LET radiation (13.3 keV/μm), the highest mutation frequencies were observed at the G1 phase. The higher LET beam always produced higher mutation frequencies in the G2/M phase than in the G1 phase, regardless of radiation dose. These results suggest that cells in the G2/M phase is hyper-sensitive for mutation induction from high LET radiation, but not to mutation induction from low LET radiation. Molecular analysis of mutation spectra showed that large deletions (which could include almost entire exons) of the mouse Hprt gene were most efficiently induced in G2/M cells irradiated with high LET radiation. These entire exon deletions were not as frequent in cells exposed to lower LET radiation. This suggests that inappropriate recombination repair might have occurred in response to condensed damage in condensed chromatin in the G2/M phase. In addition, by using a hyper-sensitive mutation detection system (GM06318-10 cells), a reverse dose-rate effect was clearly observed after exposure to carbon beams with higher LET values (66 keV/μm), but not after exposure to beams with lower LET values (13.3 keV/μm). Thus, G2/M sensitivity towards mutation induction, and the dependence on radiation LET values could both be major factors involved in the reverse dose rate effect produced by high LET radiation.

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This study examined the feasibility of applying the bubble technique to evaluate effective dose for diagnostic X-rays. A BTI-GAMMA bubble detector from Bubble Technology Industries was used for gamma detection. A multi-slab acrylic (PMMA) phantom was fabricated to quantify the effective dose E based on an ICRP-60 report. Accordingly, the bubble detectors were evaluated through preliminary tests to ascertain both the reproducibility of specific X-ray doses and the linearity of multiple X-ray doses. Qualified bubble detectors were then inserted into a multi-slab acrylic phantom. The positions of the inserted bubbles closely corresponded with the position of represented organs or tissues. The effective dose E of X-ray was determined in 12 organ and tissue samples. The bubble detector was maintained at either 21.5°C (for abdomen AP) or 22°C (for chest PA) to optimize counting, and the assessed effective doses for males and females were 66.75 ± 10.23 μSv and 66.47 ± 9.89 μSv, respectively, for each chest PA X-ray exposure. The abdominal AP X-ray exposure doses were 1183.73 ± 124.29 μSv and 976.70 ± 120.13 μSv for males and females, respectively. Controlling and holding the bubble detector at an optimal ambient temperature during X-ray exposure was the most important issue in practical application, and the optimal temperature had to be adjusted slightly with incident X-ray to effectively suppress the largest bubbles to enable easy reading.

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The effect of graded doses of high-linear energy transfer (LET) low-energy protons to induce cycle perturbations and genotoxic damage was investigated in normal human fibroblasts. Furthermore, such effects were compared with those produced by low-LET radiations. HFFF2, human primary fibroblasts were exposed to either protons (LET = 28.5 keV/μm) or X/γ-rays, and endpoints related to cell cycle kinetics and DNA damage analysed. Following both type of irradiations, unsynchronized cells suffered an inhibition to entry into S-phase for doses of 1-4 Gy and remained arrested in the G₁-phase for several days. The levels of induction of regulator proteins, such as TP53 and CDKN1A showed a clear LET-dependence. DSB induction and repair as measured by scoring for γ-H2AX foci indicated that protons, with respect to X-rays, yielded a lower number of DSBs per Gy, which showed a slower kinetics of disappearance. Such result was in agreement with the extent of MN induction in binucleated cells after X-irradiation. No significant differences between the two types of radiations were observed with the clonogenic assay, resulting anyway the slope of γ-ray curve higher than that the proton one. In conclusion, in normal human primary fibroblasts cell cycle arrest at the G₁/S transition can be triggered shortly after irradiation and maintained for several hours post-irradiation of both protons and X-rays. DNA damage produced by protons appears less amenable to be repaired and could be transformed in cytogenetic damage in the form of MN.

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The effects of ionizing radiation (IR) on tumor neovascularization are still unclear. We previously reported that vascular endothelial cells (ECs) expressing the IR-induced senescence-like (IRSL) phenotype exhibit a significant decrease in angiogenic activity in vitro. In this study, we examined the effects of the IRSL phenotype on adhesion to early endothelial progenitor cells (early EPCs). Adhesion of human peripheral blood-derived early EPCs to human umbilical vein endothelial cells (HUVECs) expressing the IRSL phenotype was evaluated by an adhesion assay under static conditions. It was revealed that the IRSL HUVECs supported significantly more adhesion of early EPCs than normal HUVECs. Expressions of ICAM-1, VCAM-1 and E-selectin were up-regulated in IRSL HUVECs. Pre-treatment of IRSL HUVECs with adhesion-blocking monoclonal antibodies against E-selectin and VCAM-1 significantly reduced early EPC adhesion to IRSL HUVECs, suggesting a potential role for the E-selectin and VCAM-1 in the adhesion between IRSL ECs and early EPCs. Therefore, the IRSL phenotype expressed in ECs may enhance neovascularization via increased homing of early EPCs. Our findings are first to implicate the complex effects of this phenotype on tumor neovascularization following irradiation.

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Heavy ion beams are a high-LET radiation that has greater biological effect than electron beams or X-rays. However, little is known about the effect of heavy ion beams on the proliferation and differentiation of human hematopoietic stem/progenitor cells (HSPCs). The present study examined the effect of heavy ion beams on gene expression in human HSPCs, especially during early stage of megakaryocytopoiesis. Human CD34⁺ cells were exposed to monoenergetic carbon-ion beams (290 MeV/nucleon, LET = 50 KeV/m) that were generated by an accelerator (Heavy Ion Medical Accelerator in Chiba). The expression of various genes related to early hematopoiesis, megakaryocytopoiesis/erythropoiesis, cytokine receptors and oxidative stress were analyzed by real-time RT-PCR. Friend leukemia virus integration 1, an early hematopoiesis-related gene, showed significantly higher mRNA expression than the control at 6 hr after irradiation. In contrast, no significant differences were observed in almost all of the other early hematopoiesis-related genes, cytokine receptor-coded genes and megakaryocytopoiesis/erythropoiesis-differentiation pathway-related genes, respectively. An analysis of the response of the genes to oxidative stress revealed the expression of heme oxygenase 1 to show a 1.5-fold and 11.9-fold increase from the day 0 control at 24 hr after 0.5 Gy and 2 Gy irradiation, respectively. Similarly, the NAD(P)H dehydrogenase-quinone 1 expression also showed a 22.0-fold and a 21.8-fold increase at 6 hr in comparison to the initial control. These results showed that the heavy ion beams affect megakaryocytopoiesis/ erythropoiesis differentiation of human HSPCs on the gene expression level.

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